Pathogenic scarring and fibrosis represent important clinical problems with potentially serious consequences for patients, including impairment of normal tissue regeneration and neighboring tissue function. Our long term goal is to identify the critical regulatory mechanisms that govern fibroblast differentiation into myofibroblasts and thereby develop new strategies to control scarring and fibrosis. Although TGF-beta is a critical component of this mechanism, current evidence suggests that both mechanical stresses and EIIIAcontaining fibronectins (FNs) are important. The EIIIA (or ED-A) segment is one of a series of spring-like segments within FN hypothesized to transduce mechanical signals. The specific aims of this proposal are to target myofibroblast differentiation by eliminating EIIIA from cellular FN in healing wounds and to identify EIIIA-dependent intracellular targets that control myofibroblast differentiation. Using EIIIA null and wild type mice, we will examine the process of fibroblast differentiation and myofibroblast function utilizing histology, immunohistochemistry, quantitative protein analyses and tensiometry. We will also test the impact of mechanical force and flanking FN sequences on EIIIA- integrin interactions, in vitro. Experiments performed with fibroblasts isolated from null or wild type mice, as well as established human lines, will determine how the EIIIA-containing FNs, relevant integrins and signaling proteins regulate myofibroblast differentiation. Findings obtained from this work will fill impotant gaps in understanding myofibroblast regulation and suggest new therapeutic strategies to modulate fibrosis.